The facile carbonized strategy is further employed to engineer the Ti-C chemical bond, serving as the touchstone. The average PL lifetime of HWT-C (4.14 ns) is much shorter than that of the 3D HWT (8.57 ns) due to the promoting effect of the chemically bonded structure on carrier separation. Consequently, the 3D HWT-C covalent photoelectrode (600 μA/cm2) exhibits a 3.6-fold increase in photocurrent density compared with the 3D HWT (167 μA/cm2). Ultimately, the model analyte of the tumor marker is detected, and the linear range is 0.02 ng/mL–100 ng/mL with a detection limitation of 0.007 ng/mL. This work provides a basic understanding of chemical bonds in tuning charge separation and insights on strategies for designing high-performance PEC sensors.The empty-space-induced depletion region in photoelectrodes severely exacerbates the recombination of electron–hole pairs, thereby reducing the photoelectrochemical (PEC) analytical performance. Herein, the chemical bond that can suppress the potential barrier and overcome the high energy barrier of out-of-plane Ohmic or Schottky contact is introduced into the PEC sensor to eliminate the depletion region and dramatically promote the separation of electron–hole pairs. Specifically, three-dimensional (3D) hierarchically wheatear-like TiO I. I3/2 anectively b were asectively . Moreoveectively . The cong) could be obtained by extrapolating the linear part of the (αhν)2 curve vs. photon energy (hν), which was obtained through the transformation of the UV-Vis DRS plot using the Kubelka–Munk function. The estimated band gaps of 3D HWT arrays and HWT-C were characterized as 3.0 and 2.39 eV, respectively.The energy band structure of 3D HWT arrays and HWT-C was also investigated in order to gain deeper insight into the promoting effect of chemical bonds on carrier charge separation. τ1 that was related to charge transfer enhanced obviously, on the contrary to the trend of τ2, which was the slow decay component related to radiative recombination [τave) was calculated using the formula of τave of the HWT-C (4.14 ns) sample was much shorter than that of 3D HWT arrays (8.57 ns), which was in accordance with the quenching trend of steady-state PL in 2. Such a conclusion was also supported by the photocurrent response, as shown in To further evaluate the facilitation of the Ti-C chemical bond on the separation efficiency of photo-induced electrons and holes, steady-state photoluminescence (PL) measurement, which was formed via the recombination of electron–hole pairs, was performed ,49. As sbination ,51,52. Tc with a correlation coefficient of 0.996. The photocurrent intensity was related to the concentration of PSA, with a range of 0.02 pg/mL to 100 ng/mL, and the detection limit (LOD) was 0.007 pg/mL. Such a larger linear range and low LOD were attributed to the Ti-C chemical bind. Furthermore, the as-constructed PEC sensor exhibited acceptable analytical performance , luteotropic hormone (LH), and human serum albumin (HSA) serving as interfering agents. The results in In this work, the chemical bond concept was first introduced into the photoelectrode of PEC bioanalysis to eliminate the depletion region between photoactive materials and promote the separation of electron–hole pairs, thereby enhancing the analytical performance. Firstly, the 3D HWT arrays were prepared with the hydrothermal method as the substrate, and the facile carbonized strategy was further employed to obtain the Ti-C chemical bond. Profiting from such robust chemically bonded interface contacts, effective spatial separation of electron–hole pairs and prominent photocurrent density were achieved, which were evidenced by PL, TRPL, and photocurrent response characterizations. Based on the larger obtained photocurrent signal, the ultrasensitive detection of PSA was realized. Exploiting the concept of the chemical bond beyond the existing strategy will offer new insights into understanding the role of the chemical bond in tuning charge separation and designing high-performance PEC bioanalysis.


Repeated anodal transcranial direct current stimulation (RA-tDCS) is a neuromodulatory technique consisting of stimulating the cerebral cortex with a weak electric anodal current in a non-invasive manner. RA-tDCS over the dorsolateral prefrontal cortex has antidepressant-like properties and improves memory both in humans and laboratory animals. However, the mechanisms of action of RA-tDCS remain poorly understood. Since adult hippocampal neurogenesis is thought to be involved in the pathophysiology of depression and memory functioning, the purpose of this work was to evaluate the impact of RA-tDCS on hippocampal neurogenesis levels in mice. RA-tDCS was applied for 20 min per day for five consecutive days over the left frontal cortex of young adult  and middle-aged  female mice. Mice received three intraperitoneal injections of bromodeoxyuridine (BrdU) on the final day of RA-tDCS. The brains were collected either 1 day or 3 weeks after the BrdU injections to quantify cell proliferation and cell survival, respectively. RA-tDCS increased hippocampal cell proliferation in young adult female mice, preferentially (but not exclusively) in the dorsal part of the dentate gyrus. However, the number of cells that survived after 3 weeks was the same in both the Sham and the tDCS groups. This was due to a lower survival rate in the tDCS group, which suppressed the beneficial effects of tDCS on cell proliferation. No modulation of cell proliferation or survival was observed in middle-aged animals. Our RA-tDCS protocol may, therefore, influence the behavior of naïve female mice, as we previously described, but its effect on the hippocampus is only transient in young adult animals. Future studies using animal models for depression in male and female mice should provide further insights into RA-tDCS detailed age- and sex-dependent effects on hippocampal neurogenesis. Neurogenesis is the complex process through which new neurons are generated and proliferated from brain stem cells, including their differentiation, before joining different final cellular networks ,2. In adTranscranial direct current stimulation (tDCS) is a safe, non-invasive neuromodulatory technique proposed as a therapeutic tool for the treatment of several psychiatric conditions, such as depression and memory deficits ,22,23,24Since adult hippocampal neurogenesis is thought to be involved in the pathophysiology of depression and memory functioning, the purpose of this work was to evaluate the impact of RA-tDCS on hippocampal neurogenesis levels in naïve female mice. Here, we hypothesize that an increase in hippocampal adult neurogenesis might contribute to the beneficial effects of RA-tDCS on mood and cognition previously observed by our team ,29. The Since neurogenesis in the hippocampus decreases significantly with age, we evaluated in the present work the effects of RA-tDCS over the left frontal lobe on cell proliferation, survival, and differentiation in the DG of the dorsal and ventral hippocampus in naïve Swiss female young adult (two-month-old) and middle-aged (ten-month-old) mice to evaluate the potential of our RA-tDCS protocol to reduce the age-induced decline of neurogenesis.+) cells, with or without RA-tDCS stimulation  nor an interaction between the hemisphere and the electrical stimulation , there was no difference between the left (stimulated) and the right  hemisphere in the number of bromodeoxyuridine (BrdUmulation . That ismulation . Data fr+ cells in naïve young adult mice (proliferation) but did not affect the number of cells that survived 3 weeks post-stimulation  = 31.92, p < 0.001) from the region  = 53.96, p < 0.001) and the stimulation × region interaction  = 5.65, p < 0.05). Although both the dorsal and ventral regions of the DG were significantly impacted by RA-tDCS, Newman–Keuls (NK) post hoc analyses revealed that the increase in cell proliferation was more robust in the dHi  than in the vHi .RA-tDCS significantly increased the number of BrdUmulation . Concern = 0.07, p = 0.79; region effect: F = 2.27, p = 0.15; stimulation × region interaction: F = 0.41, p = 0.53). The ratio of cell survival in the dHi was 75.0 ± 6.2% for the Sham group, while it dropped to 54.1 ± 2.9% for the tDCS group. In the vHi, the ratio of cell survival was 97.6 ± 9.3% for the Sham group and 70.5 ± 12.5% for the tDCS group.Concerning cell survival, RM-ANOVA did not reveal any significant effect  = 0.54, p = 0.47; region effect: F = 0.41, p = 0.53; stimulation × region interaction: F = 0.06, p = 0.80).RA-tDCS did not significantly impact cell proliferation and cell survival in the DG of the hippocampus of middle-aged (10-month-old) female mice . Concern = 1.58, p = 0.22; region effect: F = 1.85, p = 0.19; stimulation × region interaction: F = 0.10, p = 0.76). The ratio of cell survival was similar in the dHi  and in the vHi .Similarly, cell survival was not significantly impacted in middle-aged female mice  in the SVZ. They showed a bilateral increase in neuroblast numbers after cathodal stimulation and an ipsilateral (right hemisphere) increase after anodal stimulation. However, they did not observe an effect on cell proliferation in the SVZ (number of BrdU+ cells)  and mature neurons [NeuN+]) ) . These e[NeuN+]) . There aThe bilateral effects of RA-tDCS following unilateral stimulation confirm our previous data. For example, we have shown that the same tDCS protocol  bilaterally induces Zif268 expression in the basal ganglia  . FurtherWe based our study on two significant time points characterized by high  and low  rates of hippocampal neurogenesis according to Klempin and colleagues’ work ). In the+ cell numbers) and neurogenesis (BrdU+/DCX+ cells). This result was observed, to some extent, in mice of all ages, but the improvement was smaller in older mice. In fact, physical activity increased proliferation by 138% and 73% in the hippocampus of 1.5-month-old and 12-month-old female C57BL/6N mice [It is interesting to note that the increase in cell proliferation induced by tDCS in young adult mice  is not found in middle-aged mice (10-month-old), for which cell proliferation and survival rates are already very low in the Sham group. This shows that in our experimental conditions, tDCS is less effective in modulating cell proliferation when cell proliferation and survival rates are low, which is unfortunate because, from a clinical perspective, this is the time point at which we would have wanted to see the most marked effects. This age-related difference can be explained by the observation of a progressive (≈ 80%) decline in the hippocampal neurogenesis level  that has also been reported in Fischer 344 female rats  and C57B+ cells is identical in the DG of the Sham and tDCS animals at 3 weeks post-BrdU injection, tDCS animals present a significantly higher number of BrdU+ cells one day after BrdU injection. This leads to an overall higher survival rate of BrdU+ cells in the Sham group compared with the tDCS group, suggesting that an increased number of newborn neurons do not survive in the tDCS group, although we cannot draw a definitive conclusion. In middle-aged animals, the survival rate was not affected, supporting the hypothesis that tDCS was not toxic for hippocampal cells per se. This rather suggests that newly formed cells do not survive and that the effect of our RA-tDCS protocol on the hippocampus is only transient, at least in naïve female Swiss mice.Although it appears that the number of BrdUDespite the increased proliferation rates in young adults, our RA-tDCS protocol did not increase neurogenesis. Only naïve mice were used in this work (mice without depression-like behavior or cognitive impairment); therefore, we could not definitively conclude that our RA-tDCS protocol has no effect on neurogenesis levels. We can only deduce that it does not counteract an age-related decline in neurogenesis in naïve animals, but perhaps an improvement could take place if we used an animal model of depression .This will have to be performed in future studies. Indeed, David’s team reported, for example, that fluoxetine  had no effects on hippocampal neurogenesis in naïve male mice (4- to 6-month-old), whereas it significantly increased cell proliferation (+167%) and survival (+28%) in CORT-treated animals . MoreovetDCS shows potential for use in depressed patients. However, tDCS is also utilized as a cognitive enhancer in patients without depressive disorders. In fact, our team has previously reported an improvement in spatial memory in naïve 5-month-old Swiss mice . Our finExploring the effects of RA-tDCS applied at different time points should also be pursued. In fact, applying RA-tDCS two weeks after the first BrdU injections could help understand whether new cell survival rates could specifically be modulated. It would also be worth investigating whether prolonging the treatment duration  or replicating the sessions could increase the chances of modulating neurogenesis levels using our RA-tDCS protocol. Finally, it is important to emphasize that tDCS, rather than replacing AD treatments, could be used in adjunction to increase the success rate/efficacy. Future studies on neurogenesis levels in the hippocampus will be conducted in mice receiving concurrent RA-tDCS and AD treatments or RA-tDCS and physical exercise.Seventy-nine naïve Swiss female mice  were used in this study. They were group-housed  under standard laboratory conditions , except during surgery recovery and electrical stimulation periods, when they were housed individually. Food and water were available ad libitum. All experiments were conducted during the light phase of the cycle.Prior to surgery, the mice were allowed one week of acclimation to the animal facility, during which time they were repeatedly handled in order to reduce manipulation-related stress . For each experiment, the animals were divided into two experimental groups  and middle-aged mice  A.In experiment 2, cell survival and differentiation were studied in young adult  and middle-aged mice  B.All experimental procedures were performed in strict accordance with the Guide for the Care and Use of Laboratory Animals (NIH), the Animal Research: Reporting of In Vivo Experiments (ARRIVE) guidelines, and the European Union regulations on animal research (Directive 2010/63/EU). They were approved by the University of Franche-Comte Animal Care and Use Committee (CEBEA-58).® Fuji I, Leuven, Belgium; One week before starting the stimulation protocol, a tubular plastic electrode holder  was screwed into the electrode holder filled with a saline solution (0.9% NaCl). A custom-made restraining box was used to hold a larger rectangular rubber-plate electrode  was injected i.p. three times 6 h apart on the last day of the stimulation period . Given the semi-conservative mode of DNA replication, applying three injections increases the likelihood that a dividing cell will incorporate a BrdU molecule ,41.TM System  and stored at −20 °C. Coronal sections of 30 μm were obtained using a microtome , cryoprotected , and stored at −20 °C until processed. Brain sections from −0.94 mm to −3.88 mm were selected  either one day (proliferation studies) or three weeks  after the first BrdU injection. The animals were transcardially perfused first with 0.9% NaCl and then with 4% paraformaldehyde  in a 0.1 M phosphate buffer. The brains were removed, postfixed in 4% PFA overnight at 4 °C, and immersed in a 30% sucrose solution  for 24 h. The brains were then frozen using a SnapFroste bregma ). Sectioe bregma . Note th2O2 30%; Sigma-Aldrich, Saint-Quentin-Fallavier, France) in order to block endogenous peroxidase activity that could lead to non-specific background staining. DNA denaturalization was carried out by immersing the tissues in 2-N-hydrochloric acid (HCl) for 30 min at 50 °C and then rinsing in a 0.1 M borate buffer at pH 8.5. Next, the sections were exposed to donkey serum in 0.3% PBS-Triton  and then incubated overnight at 4 °C with an anti-BrdU primary antibody  diluted in PBS-T. The tissues were then incubated for 2 h at room temperature with a biotinylated anti-rat IgG secondary antibody solution . Signal amplification was conducted using an avidin horseradish peroxidase complex , and 3,3′-diaminobenzidine tetrahydrochloride  was used as the chromogen. The brain sections were then dehydrated with alcohol, cleared with xylene, and coverslipped with Canada balsam .Before staining, the selected free-floating sections were treated with a hydrogen peroxide solution , anti-NeuN , and anti-GFAP . The following secondary antibody solutions were then used at room temperature for 4 h: anti-rat IgG Cy3-conjugated , anti-mouse Alexa647-conjugated , and anti-rabbit 488-conjugated . The brain sections were then coverslipped with a glycerin + PBS-T solution (60/40%). + cells was counted bilaterally (left and right hippocampus) in the DG to assess the proliferation rate. Regarding the immunofluorescence method, images were acquired with the 20× objective of the epifluorescence microscope ApoTome.2  equipped with a digital camera  using the Axio Imager.Z2 software . The number of BrdU+ cells was counted bilaterally in the DG to assess the survival rate. In order to determine the phenotype of the surviving BrdU+ cells, co-labeled cells were counted as follows: BrdU+/NeuN+/GFAP− cells were counted as new neurons; BrdU+/NeuN−/GFAP+ cells as new astrocytes; and BrdU+/NeuN−/GFAP− cells as undetermined new cells.We analyzed an average of eight slices per animal, with four corresponding to the dorsal hippocampus (dHi) and four corresponding to the ventral hippocampus (vHi). The sections were collected at approximately the same coordinates along the entire length of the hippocampus. These coordinates were visually verified under a microscope and compared with the atlas of Paxinos and Franklin . Images +, NeuN+, or GFAP+) was measured over the entire DG and reported relative to the total size of the hippocampus studied. To obtain the number of cells in the hippocampus, the following method was used: The average cell number per slice , with stimulation  as the within-subject factor, region , hemisphere  as the between-subject factors  as the within-subject factor, region  as the between-subject factor, and the number of BrdU+ cells as the dependent variable. Newman–Keuls (NK) post hoc tests were used to describe differences between individual groups. Comparisons of the percentages for cell differentiation were assessed using a chi-square (Χ2) test (Data are presented as mean ± standard error of the mean (SEM). Significance was set at  factors , and theΧ2) test .The present study is a brief report that yielded promising results. We show that RA-tDCS modulates cell proliferation in the hippocampus of naïve young adult female Swiss mice, suggesting that unilateral electrical stimulation has bilateral physiological effects on the hippocampus. We also found, for the first time, that RA-tDCS has an age-dependent effect on cell proliferation in the hippocampus. Further experiments are needed to determine whether our RA-tDCS protocol modulates hippocampal neurogenesis in an animal model of depression and to better understand the neurobiological mechanisms involved. These additional data will help accelerate the development of this technique for use in humans to treat depression and cognitive disorders.


Of these, miR-92a-3p (AUC = 0.735), miR-221-3p (AUC = 0.774), and miR-484 (AUC = 0.725) demonstrated a predictive ability to discriminate responses from non-responses, regardless of the therapy used. Moreover, Cox regression analysis indicated that higher expression levels of miR-92a-3p (p = 0.008 **), miR-143-3p (p = 0.009 **), miR-221-3p (p = 0.016 *), and miR-486-5p (p = 0.019 *) at baseline were associated with worse overall survival, while patients expressing higher baseline miR-92a-3p (p = 0.003 **) and miR-486-5p (p = 0.003 **) had lower rates of progression-free survival. No predictive values for candidate microRNAs were found for the post-chemotherapy period. In line with these findings, we conclude that the increased baseline exosomal expression of miR-92a-3p and miR-221-3p seems to predict a lack of response to chemotherapy and lower OS. However, further prospective studies on more patients are needed before drawing practice-changing conclusions.The status of predictive biomarkers in metastatic colorectal cancer is currently underdeveloped. Our study aimed to investigate the predictive value of six circulating exosomal miRNAs derived from plasma  for chemosensitivity, resistance patterns, and survival. Thirty-one metastatic colorectal cancer patients were selected before receiving first-line irinotecan- or oxaliplatin-based chemotherapy. Blood samples were harvested at baseline and 4–6 months after the initiation of chemotherapy. The levels of exosomal expression for each miRNA were analyzed by qPCR. Our results for patients receiving first-line FOLFOX showed significantly higher baseline levels of miR-92a-3p ( Colorectal cancer (CRC) is a significant cause of cancer-related morbidity and mortality worldwide, currently ranking the fourth most diagnosed and the third most deadly neoplasia . The proThe role of biomarkers in cancer is essential not only in the initial diagnosis of the disease or at recurrence but also in the prognosis and prediction of responses to various therapies, allowing for the selection of an individualized therapeutic approach. In patients with a good performance status, the first-line treatment of metastatic colorectal cancer (mCRC) consists of chemotherapy doublets/triplets based on fluoropyrimidines, oxaliplatin, and/or irinotecan, to which a molecular targeted therapy is commonly added. Depending on the location of the primary tumor in the colon and the RAS gene’s mutational status, monoclonal anti-EGFR or anti-VEGF antibodies are chosen as appropriate . HoweverAside from the above-mentioned predictive factors for the response to therapy, there are currently no other validated clinical/biological/molecular factors for predicting the response to treatments and/or survival of these patients . IdentifMost of the miRNAs that are detectable in serum/plasma are concentrated in exosomes. Valadi and colleagues were the first team to show the transfer of miRNA through exosomes to transport genetic information . The traTo date, numerous preclinical and clinical studies have been conducted to identify and validate specific miRNAs as predictive biomarkers in metastatic colorectal cancer, yet none has been implemented for wide-scale usage in clinical practice. In this context, considering the importance of this first-line treatment option in managing patients with metastatic CRC, our study focused on evaluating the predictive role of six miRNAs of interest for assessing treatment with FOLFOX and FOLFIRI when used as a first-line therapeutic option in treating metastatic colorectal cancer.From January 2015 until December 2016, 31 patients with mCRC, who were treatment-naïve for metastatic disease, were included in this study. Baseline (B) and post-chemotherapy (PC) plasma samples (4–6 months after starting first-line chemotherapy) were available for all patients. The clinical follow-up of the patients lasted until 31 December 2019.The demographic and clinicopathological characteristics of the included patients are described in We first evaluated the signal amplification for the selected miRNAs in case–control cohorts, as follows: FOLFIRI (B), FOLFIRI (PC), FOLFOX (B), and FOLFOX (PC). Because all of the miRNAs had less than 33 CT amplification in the pooling groups , they weTo identify the associations between the miRNAs of interest and the response to first-line chemotherapy, we investigated the expression of these microRNAs in 17 patients treated with the FOLFIRI (FIRI) protocol and 14 patients with the FOLFOX (FOX) protocol at 2 different time points (baseline and post-chemotherapy). No statistically significant differences in microRNA expression between the two chemotherapy protocols were observed at baseline (B) and post-chemotherapy (PC). Our data suggest that there is no risk of associating false-positive expression levels with the response to therapy .p = 0.047 *) and miR-484  in non-responders compared to responders, regardless of the chemotherapy protocol administered  but according to the drugs used. We found increased baseline levels of exosomal miR-92a-3p , miR-146a-5p , miR-221-3p , and miR-484  expression at baseline in non-responders vs. responders , miR-484 , and miR-221-3p  had the predictive ability to discriminate non-responders from patients who exhibited a response to therapy, while the post-chemotherapy values showed suboptimal prediction accuracy  and overall survival (OS) was further evaluated. At the time of evaluation, the median PFS1 was 17 months, and the median OS was 22 months.p = 0.003 **) and miR-92a-3p (p = 0.003 **), showing that the mCRC patients with high levels of baseline exosomal expression of miR-486-5p or miR-92a-3p had significantly shorter PFS than those with low baseline exosomal levels of these miRNAs , miR-92a-3p (p = 0. 008 **), miR-221-3p (p = 0.016 *), and miR-143-3p (p = 0.009 **) is predictive for a worse OS in mCRC patients (p < 0.05) in univariate analysis were included in the multivariate analysis (MA) of survival, but none of the miRNAs showed independent predictive value for PFS1 and OS  revealed the statistically significant association of PFS1 with baseline values of miR-486-5p , and also in the subgroup of patients who received FOLFOX chemotherapy (p = 0.007). Our data indicated that high levels of miR-92a-3p at baseline in plasma exosomes are associated with low OS (p = 0.008); additionally, patients with increased expression had lower PFS1 levels (p = 0.003). These data are supported by the results of other researchers, who aimed to investigate both the prognostic and predictive roles of miR-92a-3p. In 2019, Hu’s team [n = 18) and chemo-resistant CRC (n = 18) in patients treated with FOLFOX. They showed that the level of miR-92a-3p was significantly higher in non-responders than in responders and concluded that miR-92a-3p in serum exosomes could be a good predictor of metastasis and chemoresistance in CRC. They also showed that the serum exosome level of miR-92a-3p was highest in patients with metastatic CRC vs. healthy subjects, emphasizing the diagnostic aspects and its prognostic role. Along the same lines, Fu et al. [p ˂ 0.0001), thus being both a diagnostic and a prognostic biomarker. Matsumura and colleagues [The miR-17-92p cluster has been extensively studied for its role in CRC tumorigenesis, metastasis, and responses to therapy. As a member of the miR-17-92 precursor cluster, miR-92a-3p is known to be an oncogenic miRNA in CRC. We showed that an increased baseline plasma exosome level of miR-92a-3p was associated with a lack of a therapeutic response, regardless of the chemotherapy protocol administered  and decreased miR-143-3p (p < 0.001) in CRC patients; the low expression of miR-143-3p exhibited more aggressive tumor features. Romero-Lorca and colleagues [p = 0.009). In line with our results, another clinical study [Tumor growth and metastasis are inhibited by miR-143-3p, which is reported to be a tumor suppressor in CRC. Previous studies have shown that its expression is downregulated in CRC. Sahami-Fard et al.  evaluatelleagues  includedal study  showed tp = 0.036). We did not find any studies reported in the literature that assess the role of circulating miR-146a-5p in patients with CRC. However, in a previous study, Lu et al. [Our data indicate that an increased baseline level of plasma exosomal miR-146a-5p was associated with a lack of a therapeutic response in the subgroup of patients who received FOLFOX chemotherapy  and with a lack of a therapeutic response in the subgroup of patients who received FOLFOX chemotherapy (p = 0.047). The high expression of miR-221 in CRC tissues was closely associated with a shorter survival time and a high level of risk for CRC prognosis [The results of our study show that an increased baseline expression of the oncogenic miR-221-3p in plasma exosomes was associated with low OS , but also in the subgroup of patients who received FOLFOX chemotherapy (p = 0.009). Regarding the modulation of the expression induced by chemotherapy, there was an increased expression level from baseline to post-chemotherapy of 1.68 x in responders vs. non-responders. Kjersem et al. [We pointed out that an increased baseline level of miR-484 in plasma exosomes was associated with lack of a therapeutic response, regardless of the chemotherapy protocol administered  and OS (p = 0.019) than those with low baseline expression, so it represents a negative prognostic factor in mCRC. No significant associations have been established between the level of miR-486-5p and the response to therapy. Liu and colleagues [p = 0.029) and lymph node invasion (p = 0.024). There are few data related to the role of miR-486-5p in CRC metastasis. One suggested mechanism involves miR-486-5p promoting CRC proliferation and migration by activating the PLAGL2/IGF2/β-catenin signal pathway [The last miRNA investigated in our study was miR-486-5p. We observed that patients with high levels of miR-486-5p in the plasma exosomes at baseline have shorter PFS1  and irinotecan or oxaliplatin. In most cases, chemotherapy was accompanied by a targeted therapy (anti-EGFR or anti-VEGF), according to the tumor RAS mutational status and tumor characteristics. The response to therapies was evaluated every 2–3 months by CT scans and, in only a few cases, by MRI. According to the description of the CT scans and the RECIST 1.1 criteria, patients were considered responders if they had either a complete or partial response and non-responders if they had stable or progressive disease.The whole blood for each patient was collected by venipuncture in an EDTA blood-collection tube in the same timeframe (8–12 a.m.). The samples were collected, at baseline, before the first administration of the first cycle of chemotherapy for metastatic disease, and at post-chemotherapy, 4–6 months after the initiation of chemotherapy in patients still receiving first-line chemotherapy, or before switching to second-line chemotherapy in patients who had progressed. All samples were collected before the administration of chemotherapy.Each whole-blood sample was registered under a unique code and processed immediately under similar conditions. Plasma was separated from the whole-blood samples by double successive centrifugation at 4000 and 12,000 rpm for 10 min at 4 °C. The provided plasma was aliquoted (400 µL/tube) and stored at −80 °C until its further processing.Total Exosome Isolation Kit from Plasma . An additional plasma filtration using a 0.8 μm filter was included before the precipitation step to increase the quality of vesicles specific to the exosome domain. The filtered plasma was then treated with 0.05 v/v Proteinase K and incubated for 10 min at 37 °C. After incubation, 120 µL of conditioning solution was added to the plasma, followed by 30 min at 4 °C and 10,000× g centrifugation. The resulting precipitate (plasma-derived vesicles/exosomes) was resuspended in 200 μL PBS solution and used for RNA extraction, using the Total Exosome RNA and Protein Isolation Kit  according to the manufacturer’s protocol. By introducing a supplementary step of 0.8 μm filtration, we considerably improved the exosomes’ purity by maintaining the small EVs and by eliminating the large EVs when evaluating by transmission electron microscopy (TEM) and nanoparticle tracking analysis (NTA), as previously discussed [8 transcripts/extraction) was used for each sample. A final volume of 100 μL of exosome-derived RNA was provided for each sample and stored at −80 °C until further use. The QC of the extracted RNAs was evaluated using a NanoDrop ND-1000 spectrophotometer .The isolation of plasma-derived vesicles was carried out using a specific commercial conditioning solution, iscussed . The exoThe available data regarding the predictive role of circulating exosomal miRNA for treatment responses in metastatic CRC are inadequate. Consequently, we conducted a literature study to identify new potential targets that could be investigated as predictive biomarkers for mCRC. First, we explored the specific tissue miRNAs involved in CRC development and metastasis ,39. Furt8 transcripts) as an exogenous normalizer.Because no optimal strategy is generally accepted for exosomal miRNA normalization, we choose a combination of endogenous and exogenous control miRNAs to increase the accuracy of the exosomal miRNA expression. Recent data suggest that using the same type of RNA species (miRNAs) as normalizers may be a more accurate strategy than using other species of RNA such as RNU6B small nuclear RNA (U6) . Consequ® Advanced miRNA cDNA Synthesis Kit, according to the manufacturer’s recommendations (Thermo Fisher). Compared to the classical system, the advantage of the advanced miRNA system is that it includes a pre-amplification step for the entire spectrum of miRNA contained in the sample, proportionally for all mature miRNA species existing in the samples. After the pre-amplification, each cDNA was diluted by 1/10 with RNase-free water prior to evaluation by polymerization chain reaction (PCR). Furthermore, 2.5 μL of diluted cDNA was used to investigate the expression of miRNAs of interest with a TaqMan® Fast Advanced Master Mix (2×) and specific miRNA advanced assays (Thermo Fisher Scientific) in a 10 µL reaction volume using a LightCycler  480 device under specific miR-advanced PCR conditions: activation of the UNG enzyme at 55 °C for 2 min and Taq polymerase activation at 95 °C for 20 s, followed by 40 PCR amplification cycles based on 2 amplification cycles at 3 s to 95 °C and 30 s to 60 °C, respectively. The expression level of the miRNA of interest was evaluated by ΔΔCT analysis of NR relative to R groups with the normalization of CT values at the difference between the two normalization controls, the endogenous one (miR-16-5p) and the exogenous one (cel-miR-39).For the case–control study, we generated four pools of RNA aliquots, one for each group (FOLFIRI baseline (B), FOLFIRI post-chemotherapy (PC), FOLFOX baseline (B), and FOLFOX post-chemotherapy (PC)). We chose the one-step advanced miRNA system to evaluate the level of miRNA expression, considering that it has the advantage of enabling many miRNAs to be simultaneously assessed using the same pre-amplified cDNA. A volume of 4 µL of exosome-derived RNA for each sample was used for the cDNA synthesis with the TaqMant-test for two categorical variables or the Kruskall–Wallis test, followed by Dunn’s multiple comparison post hoc test in the case of three categorical variables. The area under the ROC curve (AUC) was used to evaluate the microRNAs’ capacity to discriminate between responders and non-responders. The PFS1 was measured from the date of 1st line of chemotherapy to the date of 2nd line of chemotherapy or until the end of treatment in patients without disease progression. OS was calculated from the start of chemotherapy until the date of death or the last evaluation date. We used Cox proportional hazards regression models to evaluate the association between PFS1, respective OS, and miRNAs. For this purpose, the expression of each microRNA was dichotomized according to its median value, dividing the patients into two groups, with low and high expression.According to the data distribution, the qPCR data were evaluated with the Mann–Whitney U test or the unpaired sample Considering the small number of samples used, the data contained in this study should be regarded a hypothesis generator for future studies, given their limited statistical power for clinical applicability. Our data highlighted high baseline levels of exosomal miR-92a-3p, miR-146a-5p, miR-221-3p, and miR-484, which were correlated with lack of a response to FOLFOX chemotherapy, as well as the discriminatory power of miR-92a-3p, miR-221-3p, and miR-484 for non-responder patients regardless of the therapy used. Furthermore, increased exosomal levels of miR-92a-3p and miR-486-5p, evaluated at the onset of treatment, are associated with lower OS and PFS1, while increased exosomal levels of miR-143-3p and miR-221-3p at the beginning of therapy have a predictive value for shorter OS. Our results suggest that the increased baseline expression of exosomal miR-92a-3p and miR-221-3p predicts a lack of response to chemotherapy and a lower OS. However, the validation of these data on larger cohorts of advanced CRC patients will be able to confirm the role of these miRNAs in predicting a lack of response to FOX chemotherapy in clinical practice.


The prevalence of obesity in general pediatric population increases without sparing children with T1D. We intended to find factors associated with the possibility of preserving endogenous insulin secretion in individuals with long-standing T1D. At onset, higher BMI is associated with higher C-peptide level, which may indicate to be one of the favorable factors involved in preserving residual β-cell function. The study determines the influence of BMI on C-peptide secretion in children newly diagnosed with T1D in two years observation.We assessed the possible relationship between selected pro- and anti-inflammatory cytokines, body mass at recognition and β-cell function status. 153 pediatric patients with newly diagnosed T1D were divided into quartiles according to BMI-SDS index. We separated a group consisted of patients with BMI-SDS >1. Participants were followed up for two years and examined for changes in body weight, HbA1c, and insulin requirement. C-peptide was assessed at baseline and after two years. We evaluated the patients’ levels of selected inflammatory cytokines at baseline.Subjects with higher BMI-SDS presented higher serum C-peptide levels and lower insulin requirements at diagnosis than children with lower body weight. The two-year follow-up showed that C-peptide levels of obese patients dropped more rapidly than in children with BMI-SDS within normal limits. The group with BMI-SDS >1 showed the greatest decrease in C-peptide level. Despite statistically insignificant differences in HbA1c at diagnosis between the study groups, in the fourth quartile and BMI-SDS >1 groups, HbA1c as well as insulin requirements increased after two years. The levels of cytokines varied the most between BMI-SDS <1 and BMI-SDS >1 groups and were significantly higher within BMI-SDS >1 group.Higher BMI, associated with enhanced levels of inflammatory cytokines, relates to preservation of C-peptide at T1D recognition in children but is not beneficial in the long term. A decrease in C-peptide levels combined with an increase in insulin requirements and in HbA1c among patients with high BMI occur, which may indicate a negative effect of excessive body weight on the long term preservation of residual β-cell function. The process seems to be mediated by inflammatory cytokines. HoweverMoreover, C-peptide levels presented in some studies as an independent factor of partial remission and is correlated with its duration , 14. TheNevertheless, C-peptide levels are firmly and independently related to body weight and age at and shortly after the diagnosis of T1D . There aIntriguingly, some studies have shown that higher BMI is related to elevated levels of pro-inflammatory cytokines, which could potentially affect the destruction of pancreatic islets –29. The In our study we hypothesized that greater BMI is associated with higher C-peptide level at diagnosis, but during follow-up C-peptide levels dramatically decrease along with reduced residual pancreatic β-cell function. However, few studies evaluating the effects of obesity-associated inflammatory cytokines on residual β-cell function have been done so far, thus we decided to investigate this relationship.2Our prospective observational research covered 153 youths newly diagnosed with T1D in the Department of Pediatrics, Endocrinology, and Diabetology with Cardiology Division, Medical University of Bialystok, Poland, who were consistently examined trimonthly in Outpatient Clinic during two years. Every patient received insulin intravenously at the time of diagnosis. The therapy has been changed to functional intensive therapy (multiple daily injections (MDIs) or continuous subcutaneous insulin infusion (CSII)) as the method of choice when the patient’s condition was stabilized. The main inclusion criteria were ISPAD diagnosis criteria of type 1 diabetes and age under 18 years. Exclusion criteria were diagnosis of other types of diabetes, such as MODY and type 2 diabetes. The patients were encouraged to maintain their normal physical activity in accordance with ISPAD recommendations .Anthropometric data such as gender, height, weight and body mass index (BMI) have been assessed. To adjust for age and gender, the BMI SD score (BMI-SDS) was calculated using sex-specific and age-specific BMI growth charts based on local Polish OLAF study . All theDIR mean values were calculated depending on the information collected during the medical appointments from the course of previous 2 weeks, from patient’s home monitoring diary or personal insulin pump reports. Total DIR and HbA1c at the disease onset, as well as one year and two years after diagnosis were evaluated. The analyzed results of fasting C-peptide levels on the first admission to hospital and after two-years of observation are given as [ng/mL] and a cut-off level of clinical validity was adjusted from 0.2 nmol/L to 0.6 ng/mL .Levels of cytokines were determined once, at disease onset as previously described and published . Plasma After the patients had recovered from ketoacidosis, CRP and morphology tests were performed as part of standard clinical procedure along with other laboratory tests. The results were within the normal range and precluded a clinical diagnosis of inflammation at that point.3post-hoc RIR test for unequal numbers or the ANOVA rang Kruskal-Wallis test and the median test with post-hoc tests of multiple comparisons. All data are expressed as mean ± SD. Relative risk was evaluated with the use of Fishers’ exact test, and calculation of 95% confidence interval with Koopman asymptomatic score. We performed Pearson correlation analysis to analyse possible associations of intensity of pancreatic autoimmunity  and BMI, BMI-SDS, HbA1c, DIR and C-peptide. We also performed an analysis of covariance (ANCOVA) including gender and age as the accompanying variables to verify their possible influence on the ANOVA results between groups stratified by BMI-SDS into quartiles.Statistical analysis was performed using Statistica version 13.3 . The Kolmogorov-Smirnov test of normality was used to test the distribution of all continuous variables with Lillefors correction and Shapiro-Wilk tests. All examined variables had a parametric distribution and were evaluated using the unpaired Student t-test to compare the differences between the groups and as several of the studied parameters were not normally distributed, the two-tailed Mann-Whitney U-test was used to correlate continuous variables. In the analysis of more than two groups, the analysis of variance was used with Tukey’s Differences were considered statistically significant with p-value below 0.05. Significant changes were demonstrated within the graphs or tables with asterisks or exact p-value: * - p < 0.05, ** - p < 0.01, *** - p < 0.001, **** - p < 0.0001.4We obtained approval of the Ethical Committee at the Medical University of Bialystok (R-I-002/435/2017). Written informed consent was obtained from parents/legal guardians as well as from patients aged > 16 years.5The study included 153 Caucasian children with a proportional ratio of both sexes (49% boys). The clinical and biochemical characteristics of the study group are collected in We found that average HbA1c at the first admission to the hospital among all the groups was comparable, with mean value 11.46 ± 2.65%. Nevertheless, after two years there were noticeable differences in HbA1c levels, especially in comparison of first and fourth BMI-SDS quartile groups . We have also observed a slight difference in HbA1c in BMI-SDS<1 an BMI-SDS>1 groups   2.DIR of the study groups at hospital discharge and two years after diagnosis was also compared Table 2.We also observed that during one year of follow-up, between the first and second year of observation, HbA1c and DIR increased in the subgroup of children within BMI-SDS>1. HbA1c level increased from 6.53% to 7.02% (P = .152) and DIR from 0.55 to 0.75 U/kg/24h (P = .045). It should be added that this group of children maintained their weight throughout the observation period .Next, we analyzed fasting C-peptide secretion in our patients at admission and two years after diagnosis Table 2.Eventually, we measured patients’ levels of inflammatory cytokines such as IL-1β, IL-10, IL-12p70, IL-17, TNF-alpha and IFN-gamma at the time of diagnosis Figure 2Our analyses show that groups with BMI-SDS>1 and BMI-SDS<1 did not statistically differ in age (P = 0.71), pH (P = 0.23) and base excess (P = 0.1) at the time of diagnosis. Moreover, we observed that differences between age and pH at admission between the first and fourth quartiles were not substantial . All the mean values of the BMI-SDS <1 and >1 groups characteristics including parameters affecting C-peptide level as well as their comparison with respect to the P value between groups at the disease onset, after one year and two years of observation are presented within Correlation analysis adjusted for age considering the possible associations of intensity of pancreatic autoimmunity  with BMI-SDS, BMI, HbA1c and C-peptide showed no statistical significance between studied autoimmunity and other variables within the entire study group. However, in correlation analysis adjusted for age performed between BMI-SDS at onset and inflammatory cytokines we found significant correlation with IL-10 (r = .316 P = .033) as well as with IL-17 . Additionally, BMI-SDS correlated significantly with C-peptide at onset , DIR at discharge  and HbA1c after 2 years  Table 3.th quartile group , in the group above the median BMI-SDS , and similar trend for the BMI-SDS>1 .Interestingly, there are some statistically significant correlations between GADA and BMI-SDS at onset within particular sub-groups: in the BMI-SDS 4In our final analysis, we intended to evaluate predictive potential of selected diabetes-related parameters in establishing outcome of the therapy. First, we observed that patients with values of c-peptide lower than 0.7 ng/ml at the admission stage had significantly higher risk of keeping that parameter below clinically significant level after 2-year treatment (p = 0.0125). On the other hand, the same group of pediatric subjects had slightly higher tendency for maintaining normal level of SDS-BMI (p = 0.0862). As presumed, those patients who demonstrated initially SDS-BMI values of above 1, had substantially higher risk of that parameter elevated also after 2 years of therapy (p < 0.0001). Similarly, for subjects with lower median HbA1c levels (below 11%) at admission, the odds for these patients for reaching HbA1c normalization were significantly increased (p = 0.0163) Figure 36The current study was designed to determine the impact of BMI on the preservation of residual β-cell function in children with newly diagnosed type 1 diabetes. We found a positive correlation between BMI-SDS and serum C-peptide level in our patients, occurring only at the time of diagnosis. Interestingly, after two years of follow-up, we observed a sharp decrease in serum C-peptide level, as well as an increase in daily insulin requirement and HbA1c in the group of children with the highest BMI-SDS, indicating that in long-term observation higher BMI affects residual β-cell function negatively. Previous studies support our finding that at the time of diagnosis, children with higher BMI-SDS demonstrate more preserved β-cell function than thinner ones. Szypowska et al. in their study revealed that obese and overweight children show higher fasting C-peptide levels as well as lower HbA1c levels compared to leaner individuals at the baseline . In addiIn contrast to the studies mentioned above, our study includes a two years follow-up of the subjects. Additionally, consistent to our observations is similar Swedish national study conducted by Ludvigsson et al., who showed in their one-year follow-up of children with newly diagnosed diabetes that subjects with higher BMI presented higher levels of C-peptide at diagnosis, whereas they lost more C-peptide during the first year after the onset of the disease than those with lower BMI . On the Our observation showed that the third quartile BMI-SDS group experienced the smallest decrease in C-peptide level during the two-year follow-up, which may indicate that children with such body mass (mean BMI-SDS -0.72 ± 0.29) preserve β-cell function the most over the progression of type 1 diabetes. It is important to note that the first quartile BMI-SDS group showed almost the same decrease in C-peptide levels as the SDS-BMI >1 group which means that the leanest children do not preserve the function of the pancreatic islets in the best manner.Furthermore, we found that children with BMI-SDS>1 were close to achieving criteria for PR after one year of implementing intensive insulin therapy, whereas two years after diagnosis their insulin requirements significantly increased, which may indicate that high BMI is not a factor that can contribute to prolonged PR period. In our relative risk analysis children with SDS-BMI above 1 occurred unfortunately to maintain increased body mass during the two years observation. This situation opens new therapeutic strategies – to emphasize the great need of normalization of body mass in those T1DM recognized with overweight. The negative effect of BMI on the preservation of pancreatic β-cell function in children with type 1 diabetes may be potentially explained by Wilkin’s accelerator hypothesis according to which the differences between type 1 diabetes and type 2 are blurred both clinically and etiologically . TherefoInterestingly, some studies show a positive correlation between BMI and levels of inflammatory cytokines , 26, 44.via autoimmune mechanisms (On the other hand, the correlation we have observed between BMI-SDS and GADA appears to confirm the destructive impact of obesity and its concominant low-grade inflammation on pancreatic beta cells chanisms , 48. Ourchanisms .+TIGIT+CD8+ memory T cells, which show reduced IFNγ and TNFα secretion, which may indicate their functional exhaustion (The presence of a higher residual beta cell mass in individuals with type 1 diabetes could provide an opportunity to propose therapeutic interventions targeting the immune system in these patients. In development or under testing are several therapies aimed at modifying the immune system’s response to preserve beta cell function. Teplizumab, a CD3-directed monoclonal antibody was approved in November 2022 in the USA as a first drug indicated to delay the onset of stage 3 T1D in adults and pediatric patients aged 8 years and older with stage 2 T1D . Recent haustion . Likewishaustion . It was haustion .It should be added that the higher, unfavorable inflammatory milieu observed in patients with higher BMI may also reduce the efficiency of therapeutic interventions. It is proven that inflammation contributes to the development of insulin resistance , thus inNevertheless, our study has some limitations. The study was performed in a single center, thus our group of patients was not large and diversified enough. Moreover, only 22 children were classified into the BMI-SDS >1 category, since the BMI-SDS <1 consisted of 121, the results might have been affected by statistical errors. We believe that also the correlation of cytokines and C-peptide levels do not show statistical significance because of the small study group. Therefore, a wider range of further research and epidemiologic studies with a greater number of varied patients are required. Additionally, the influence of factors that may affect C-peptide levels, such as history of infections and puberty , 55 was 7Higher BMI at type 1 diabetes onset in children, connected initially with preserved C-peptide, does not protect from rapid decrease in β-cell function in these patients. Significant decrease in residual β-cell function within two years from diagnosis in a group of children with the highest BMI may be the result of negative impact of elevated levels of inflammatory cytokines on pancreatic β-cells in the course of obesity. In terms of C-peptide preservation, it is most beneficial for type 1 diabetes sufferers to keep their BMI-SDS similar to individuals in the third quartile group of our follow-up, and it should be noted that neither the highest BMI-SDS nor the lowest BMI-SDS have a beneficial effect on preserving residual beta-cell function. Moreover, higher BMI is not associated with a greater likelihood of prolonged PR period. Since it has been proven that residual β-cell function has an advantageous result on metabolic control several years after disease onset this issue needs further investigation and prospective studies. Therapeutic strategies to maintain body mass within third centile of BMI-SDS ranges should be developed in order to prolong C-peptide, hence endogenous insulin secretion in new onset type 1 diabetes in children.The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.The studies involving human participants were reviewed and approved by Ethical Committee at the Medical University of Bialystok. Written informed consent to participate in this study was provided by the participants’ legal guardian/next of kin.Conceptualization, EK, SC and BG-O. Data curation, EK, SC, MJ-S and BG-O. Methodology, EK, SC, MJ-S, AP, AS and KG. Formal analysis, EK, SC, AS and KG. Investigation, EK, SC and MJ-S. Resources, EK, SC, MJ-S, AP and BG-O. Writing – original draft, EK and SC. Writing – review & editing, KG, MM, AB and BG-O. Visualization, BG-O. Validation, MM, AB and BG-O. Project administration, BG-O. Supervision, MM, AB and BG-O. Funding acquisition, BG-O. All authors contributed to the article and approved the submitted version.


Lung cancer is a common malignant tumor, which is seriously harmful to human life and health. Nowadays, it has gradually become one of the best treatments for non-small cell lung cancer (NSCLC) to combine immunotherapy and chemotherapy, and its clinical efficacy is preliminary. Nevertheless, substantial differences exist between various studies and various indicators. Despite their unconvincing results, high-quality research evidence is needed to support them. In this case, further correlative studies are necessary to investigate the prognostic outcomes of PD-1/PD-L1 suppressors in combination with chemotherapeutic drugs in NSCLC.The online public databases were searchable for the clinical trials that consisted of NSCLC patients who had concluded their chemotherapy and who had accepted PD-1/PD-L1 suppressors. The time-span of the search spanned from the beginning to the end of the database. Two investigators retrieved the data independently. RevMan 5.3 statistical software was utilized for the assessment of bias risk. The software followed the Cochrane Handbook 5.3 guidelines.There were seven clinically controlled studies with 2781 NSCLC samples finally included in this study. A meta-analysis of the post-treatment overall response rate (ORR) was undertaken. A remarkably higher ORR rate was observed in the study group (p<0.05). Study participants had a noticeably longer PFS . Study participants had markedly longer overall survival (OS) . The incidence of adverse events (AEs) of Grade 3 or above was not clinically clearly different (P>0.05), as demonstrated by the incidence of AEs. The funnel plots were separately charted in accordance with ORR rate, PFE, OS, and Grade 3 AEs. The majority of the funnel plots were symmetrical and a minority of funnel plots were asymmetrical, indicating the heterogeneity of research and the limited evidence available may lead to some publication bias in the contained literature.The combined PD-1/PD-L1 inhibitors with conventional chemotherapy can dramatically elevate the prognosis of NSCLC patients, obviously enhancing the ORR rate and prolonging their PFS and OS. Furthermore, it was found that adding PD-1/PD-L1 inhibitors to conventional chemotherapy did not result in any additional adverse effects. In accordance with the latest research statistics of the International Agency for Research on Cancer (IARC) of the World Health Organization, there will be 82000 new cases of lung cancer in China happened in 2020, with 71000 deaths. Taking into account their histology, in addition to small cell lung cancer (SCLC), there is also non-small cell lung cancer (NSCLC), and the latter account for 85% of lung cancers . The tarImmune checkpoint inhibitor (ICI) drugs are capable of inhibiting the “off-work break-feed” signal by neoplastic cells , which c22.1The design was developed based on the preferred reporting items of the system review and meta-analysis plan. To comprehensively understand the effect of PD-1/PD-L1 inhibitor combined with chemotherapy drugs on the prognosis of non-small cell lung cancer through systematic review and meta-analysis.2.2All cases undergo pathological examination. The pathological type was NSCLC without EGFR, ALK and other driving gene mutations.2.3PD-1/PD-L1 inhibitors  were administered along with standard chemotherapy to the study group, whereas standard chemotherapy or placebo was given to the control group.2.4Compare the ORR, PFS and OS indicators of the included patients.2.52.5.1(1) Study types: Clinical trials that did not involve PD-1/PD-L1 inhibitors and chemotherapy. Additionally, references were retroactively included in the literature to supplement and obtain relevant information. If the results of the same clinical control study were published in a series of articles, we contained only the latest and most comprehensive literature (2). Design type: clinical control study.2.5.2(1) Clinical control studies were not conducted (2); As a result of the incomplete report, it was not possible to use the data (3); Recent studies were taken into consideration, and a repeat of the study’s content was conducted (4); there were no noticeable effects of the study in terms of curing. (5) studies that did not provide access to the full-text studies repeatedly published through multiple channels and studies in languages other than Chinese or English.2.6Data about advanced NSCLC patients treated with PD-1/PD-L1-free inhibitors associated with chemotherapy were collected from PubMed, EMBASE, ScienceDirect, Cochrane Library, China National Knowledge Infrastructure (CNKI), VIP full-text (VIP), Wanfang Database and Chinese Biomedical Literature data (CBM), conference papers, degree papers. In addition to free words, subject words were searched, and key words included PD-1/PD-L1 inhibitors, chemotherapy, and patients with NSCLC; short-term efficacy; prognosis; meta-analysis; advanced prognosis from January 2010 to March 2022.2.7The included references are imported into EndNote 20, and the repeatedly selected documents are eliminated through EndNote 20. Manual screening based on literature content. Next, two reviewers screen the study based on inclusion and exclusion criteria. The work of the two reviewers was carried out independently, and the reasons for the exclusion of the literature were recorded. The documents with differences are discussed by the third reviewer. The final step is to check whether there is any research overlap in all the included literature.2.81) The study contained an assessment of bias risk. The bias risk estimate was performed with ReviewMan 5.4 based on Cochrane Handbook 5.3.2) Literature screening and data extraction. A cross-check was conducted by two researchers on the results who conducted independent screenings of literature, gathered data, analyzed quality, and assessed quality. In case of lack of complete data in the literature, the authors of this paper would be contacted. Besides basic information about the authors, publication date and number of cases, the data extraction included information about interventions, therapeutic procedures and outcomes.2.9via inverted funnel plots. The asymmetry of the funnel plots was examined using Eggers’ test. For tests with P-values less than 0.1, we employed the Trimand Fill method to rectify for potential release bias.RevMan5.3 software was derived from the Cochrane Collaboration Network Meta-analysis. Odds risk (OR) was adopted as an indicator, and the measured data were presented as mean difference (MD). Every effect quantity was calculated by using point estimates and 95% confidence intervals (CI). Heterogeneity was examined using χ 2 for heterogeneity and I2 was used to determine heterogeneity. If P>0.05 and I2<50%, it was considered homogeneous and the modified effect model was acquired for Meta-analysis; if P<0. 05 and I2≥50%, the combined effect was needed to determine homogeneity and a random-effects model was selected. During P<0.05, the original heterogeneities would not be chosen, descriptive analysis was applied. It was statistically remarkable that the differences existed (p<0.05). In addition, literature bias in publications was determined 33.1Through computer database retrieval, 7 clinical controlled studies were finally contained –17, with3.2In this meta-analysis, all seven clinical controlled studies reported baseline patient status. A total of four studies didn’t describe the random method in detail and discussed detailed treatment measures that mentioned “random distribution” in the literature. It was found in the seven literature that there was a high number of blind methods as well as lost follow-up or withdrawals with detailed descriptions. The improved JADAD score of 7 articles was 3 or 4, all of which were of high quality. 3.33.3.12 = 0.83, df=2, P=0.66, I2 = 0%, indicating that a distinct heterogeneity could be observed in the study’s data. Using a fixed effect model analysis, study participants had a higher ORR rate . In combination with chemotherapy, PD-1/PD-L1 inhibitors noticeably extended the survival of patients with NSCLC (The OS after treatment was analyzed in a meta-analysis. Heterogeneity analysis results indicated that Chith NSCLC Figure 63.3.52 = 23.26, df=6, P=0.0007, I2 = 74%. It indicated that a distinct heterogeneity could be observed in the study’s data. Using the random effect model, no noticeable differences were found in adverse reaction incidences (P>0.05, Clinically, AEs above Grade 3 required special intervention or face dose reduction or even drug withdrawal. Meta-analysis was conducted on AEs above Grade 3. Heterogeneity analysis results indicated that Chi P>0.05, Figure 73.3.6Funnel charts were drawn on the basis of ORR rate, PFE, OS and Grade 3 AEs respectively. The publication bias analysis was conducted 11. Ther4Cancer-related deaths account for roughly 20% of all deaths caused by lung carcinoma, which represents one of the clearest malignancies . The incImmunotherapy is by far of the most promising strategies . Agents It is noteworthy that NSCLC patients can have a better survival time after ICI treatment, but some patients still fail to benefit. Additionally, NSCLC patients receiving ICI also experience immune-related adverse effects that are associated with improved survival. It was found that ICI combined with chemotherapy has good OS and PFS, and the treatment effect is better than ICI alone. PD-1/PD-L1 inhibitors are better controlled by glucocorticoid therapy even if adverse reactions can occur, and have a better prospect of application .In the present study, 2781 cases were sampled from seven controlled clinical trials. In the study group, there was a marked improvement in ORR, indicating that chemotherapy combined with PD-1/PD-L1 inhibitors can remarkably enhance the efficiency of patients with NSCLC with remarkable short-term effects. PD-1 is a human immunoglobulin that specially binds to human PD-1, allowing PD-1 to interact with PD-L1 and PD-L2 ligands. In addition to achieving desirable therapeutic effects, it has outstanding clinical advantages as a targeted cancer treatment agent. Through the employment of PD-1/PD-L1 inhibitors in combination with conventional chemotherapy, further improvements in clinical outcomes can be achieved. In the meta-analysis, PFS and OS were remarkably prolonged after treatment in the study group. Combined treatment with PD-1/PD-L1 inhibitors on top of chemotherapy remarkably prolonged progression-free survival and OS in NSCLC patients and could remarkably improve the long-term prognosis of patients. Chemotherapy combined with PD-1/PD-L1 inhibitors promotes remarkable antitumor effects and prolongs patients’ survival.Clinical management of AEs associated with combination chemotherapy with PD-1/PD-L1 inhibitors has proven to be an essential aspect of therapy , 38. The5When combined with conventional chemotherapy, PD-1/PD-L1 inhibitors can noticeably enhance the outcome of NSCLC patients, noticeably increasing the ORR rate and prolonging their PFS and OS. Furthermore, a combination of PD-1/PD-L1 inhibitors and conventional chemotherapy did not appear to increase AEs.The datasets used and/or analysed during the current study are available from the corresponding author on reasonable request.MS were major contributors in writing the manuscript. CL collected the patient data. JG did literature searches, and revised the manuscript. All authors contributed to the article and approved the submitted version.


Post-operative systemic inflammation response syndrome (SIRS) is an event that results from surgical trauma, white blood cells contact activation, and intra-surgical bacterial translocation, which is difficult to distinguish from sepsis. Presepsin is a novel biomarker that is increased since the early stages of bacterial infection and can be used to confirm the diagnosis of post-operative infectious complications. This study aimed to investigate the diagnostic performance of presepsin for post-operative infectious complications compared to other well-known biomarkers.This cross-sectional study included 100 post-operative patients admitted to Cipto Mangunkusumo National Hospital and Bunda Hospital in Jakarta, Indonesia. The objective was to identify the optimal cutoff and trend of plasma presepsin concentration on the first and third day after surgery and to compare them with other biomarkers.Plasma presepsin level was higher in the infection group compared to the non-infection group . Presepsin levels tended to increase on the third post-operative day (median + 252 pg/ml) in children with infection. The opposite trend was observed in the non-infection group from the first to the third day (median -222.5 pg/ml). Presepsin delta, a three-day difference between the first and third post-operative day, had the best diagnostic performance compared to other biomarkers (Area Under the Curve 0.825). The optimal cutoff for presepsin delta to diagnose post-operative infection was +90.5 pg/ml.Serial assessments of presepsin levels on the first and third days post-surgery and their trends are helpful diagnostic markers for clinicians to detect post-operative infectious complications in children. This condition is frequently confused with sepsis . MicrobiIn the past decade, presepsin (sCD14-ST) has been studied as a novel marker of infection. It is a biological immune marker with a potential role in detecting infection, especially in its early stages –8. PlasmAt the time of this study, only one systematic review and meta-analysis had been published regarding the diagnostic performance of presepsin in pediatric sepsis. It reported better performance of presepsin (Area Under the Curve/AUC 0.94) compared to CRP (0.51) and PCT (0.76) in detecting sepsis in children . There h2.2.1.This cross-sectional study was conducted on children aged 1 month-18 years in Cipto Mangunkusumo National Hospital and Bunda Hospital in Jakarta, Indonesia. The Ethics Committee of the Faculty of Medicine, Universitas Indonesia, approved this study. Samples were collected prospectively through consecutive sampling from January to November 2021. Sample size was estimated by using the sample size calculation formula for diagnostic test . The inc2.2.Complete blood count, plasma presepsin and procalcitonin levels, and blood culture samples were collected within 24 h after surgery. The second presepsin and procalcitonin sample was collected on the third day after surgery. Other microbiological cultures were also obtained once there were signs of infection.TM with hydrodynamic focusing method, flow cytometry method using a semiconductor laser, and sodium lauryl sulfate-hemoglobin method. Presepsin was measured using the PATHFASTTM immunoanalyzer . The blood culture was analyzed using BACTECTM with BacT/Alert method .Complete blood count was performed using Sysmex XT 2000i2.3.p < 0.05.Data analysis was conducted with the 26th version of the Statistical Package for the Social Sciences (SPSS) statistical program. Receiver operating characteristic (ROC) analysis was used to determine the presepsin AUC and cutoff value to diagnose post-operative infection, and the optimal cutoff value was chosen using Youden's index. Furthermore, the sensitivity, specificity, negative predictive value (NPV), positive predictive value (PPV), and likelihood ratio (LR) were calculated following the ROC curves for each biomarker. Statistical significance was set at 3.3.1.One hundred post-operative subjects as summarized in n = 5) occurred in multiple organs. Twenty-seven bacteria and 2 fungi were cultured from 18 patients. Most infections were caused by Gram-negative bacteria, such as Escherichia coli and Klebsiella pneumoniae with 8 and 7 cultures, respectively.The prevalence of post-operative infection was 18 of 100 subjects (18%). One-third of the infection (3.2.p < 0.05).The difference between the first and third-day presepsin levels was calculated as “delta presepsin.” This variable was determined to evaluate the trend of presepsin levels on the first and third day after surgery. p = 0.418).The ROC curve showed optimal cutoff for third-day presepsin, delta presepsin, first-day PCT, and third-day PCT were 761 pg/ml, + 90.5 pg/ml, 0.73 mg/L, and 1.085 mg/L, respectively. 4.In this study, post-operative infection was prevalent in 18% of patients. Specific data on children remain scarce, and most are focused on surgical site infection (SSI). These findings are comparatively higher than those of other studies –16, whicThe median first-day and third-day presepsin levels in this the current study were 725 pg/ml and 531.5 pg/ml, respectively. This result was higher than the average presepsin level in healthy adult subjects, i.e., 89–382 pg/ml . StudiesThe median post-operative presepsin levels on the first and third days in the non-infection group, respectively, were 717 and 516 pg/ml. This result was interesting as the non-infection group results were similar to the median presepsin level to detect sepsis in other studies. A systematic review by Yoon et al. reported a 650 pg/ml cutoff with 0.983 AUC to detect sepsis in children . Higher . on post-esophagostomy surgery subjects reported a median first- and third-day post-operative presepsin levels of 313 pg/ml (IQR/interquartile range 218–398) and 276 pg/ml (IQR 229–361), respectively, in the non-infection group , CRP (0.8 AUC), and neutrophil-lymphocyte ratio (0.804 AUC) . AccordiIn this study, first- and third-day PCT level were significantly increased in the post-operative infection group. However, it was notable that PCT increased similarly from first to third day post-surgery in both infection and non-infection groups. This is in contrast with presepsin, which predictably decreased on the third day post-surgery in the non-infection group, and increased on the third day post-surgery in the infection group. The cut-off levels were also considerably lower compared to other studies . Farias et al. reported 12.9 ng/ml cut-off in the infection group compared to 5.6 ng/ml in the non-infection group on the first day post-cardiac surgery; and 15 ng/ml (infection group) compared to 3.6 ng/ml (non-infection group) on the third day post-cardiac surgery . The difThis study showed that PCT was an effective parameter for infection especially within the first 24 h. This is highly beneficial in daily practice, as it is crucial to determine infection as soon as possible in order to start the appropriate antibiotic therapy. However, empirical antibiotic therapy was started on the first post-operative day in our samples, as all of the subjects underwent major surgeries. In such cases, presepsin may help clinicians not only to evaluate the possibility of infections; but also the response to the current antibiotic therapy. The alternate presepsin level will help clinicians to differentiate infection from clinical features of SIRS in post-operative patients. Increased presepsin levels on the third post-operative day help clinician to confirm the presence of infection and promptly implement the right course of action, such as changing the antibiotics, or increasing the dose to a dose appropriate for severe infection.This study is the first to document post-operative presepsin levels and changes in children within the first 72 h following surgery. Furthermore, subjects with a lower preoperative glomerular filtration rate were excluded because this condition could increase plasma presepsin levels through decreased excretion , 20.A limitation of this study lies in the fact that only presepsin levels on the first and third days after surgery were evaluated. The operational definition of post-operative infection is an infection occurring within 30 days post-surgery or 90 days for several specific procedures or implants . There wThis study evaluated blood cultures on the first post-operative day. Another limitation was that the next cultural examination was conducted based on the specialist's consideration, which may vary for each subject. We also did not collect plasma presepsin and PCT along with the culture examination beyond the third post-operative day. Ideally, the plasma presepsin and PCT should be taken simultaneously together with every culture samples.5.Both presepsin and PCT can be used as diagnostic markers in predicting post-operative infection in children. Delta presepsin (difference between first- and third-day presepsin levels) had the best diagnostic performance compared to other biomarkers.


Real world data on the response to the SARS-CoV-2 vaccine in patients with immunomediated diseases (IMIDs) treated with immunesuppressants is of great interest because vaccine response may be impaired. The main aim was to study the humoral and cellular immune response after SARS-CoV-2 vaccination in patients with IMIDs treated with immunosuppressants. The secondary aim was to describe the frequency of SARS-CoV-2 infections after vaccination in these patients.This is an observational study including 86 patients with IMIDs. All patients were treated with biologic or targeted synthetic disease-modifying antirheumatic drugs . Demographic and clinical information were collected. After 4–6 weeks of 2nd and 3rd vaccine doses, humoral response was assessed using the Thermo Scientific ELiA SARS-CoV-2-Sp1 IgG Test. Also, in patients with serum SARS-CoV-2 antibody levels under 100UI/ml, cellular response was analyzed using the QuantiFERON SARS-CoV-2 Starter Pack.A total of 86 patients under b/tsDMARDs and 38 healthy controls were included. Most patients received TNFi . SARS-CoV-2 antibodies (Ab) were present in an 86% of patients with IMIDs and in 100% healthy controls (p = 0.017). However, 12 (14%) patients had undetectable SARS-CoV-2 Ab levels, all treated with rituximab. In addition, SARS-CoV-2 Ab (IU/ml) were statistically lower in patients (Mdn (IQR): 59.5 (17–163) in patients vs 625 (405–932) in controls, p < 0.001). Patients treated with rituximab had lower Ab levels than those treated with TNFi and controls (p < 0.001). The cellular response to SARS-CoV-2 vaccine was evaluated in 30 patients. Eleven patients had a positive cellular response, being more frequent in patients treated with rituximab (p = 0.03). SARS-CoV-2 infection was reported in 43% of patients and 34% of controls after vaccination. Only 6 (7%) patients required hospitalization, most of whom treated with rituximab (67%).SARS-CoV-2 antibody levels were lower in patients than in controls, especially in patients treated with rituximab. A cellular response can be detected despite having a poor humoral response. Severe infections in vaccinated patients with IMIDs are rare, and are observed mainly in patients treated with rituximab.The online version contains supplementary material available at 10.1186/s13223-023-00832-0. The SARS-CoV-2 pandemic resulted in high morbidity and mortality due to COVID-19. The risk factors associated with worse COVID-19 outcomes among the general population include older age, sex, and chronic diseases . EvidencVaccination strategies protect against infections through stimulation of the humoral and cellular immune responses. The relative importance of humoral and cellular immunity in conferring protection from infection varies with the individual microorganism . AntibodVaccination against SARS-CoV-2 has been the largest vaccination program in the history of the Spanish National Health System, and immunocompromised patients have been prioritized in vaccination schedules. The vaccines approved for prevention of SARS-CoV-2 infections include mRNA vaccines , non-replicating viral vector vaccines , and traditional inactivated whole virus vaccines . All currently available vaccines against SARS-CoV-2 have proven to be effective in clinical trials. However, their efficacy has not been tested in immunocompromised patients . A recenImmunosuppressive therapy plays a leading role in the impaired response in patients with IMIDs. Conventional, biologic, and targeted synthetic disease-modifying antirheumatic drugs (c/b/ts-DMARDs) and corticosteroids are widely used in monotherapy or in combination to treat affected patients. Some of these drugs have been shown to impair the response to vaccines . Data onAlthough there is increasing evidence on how the response to the SARS-CoV-2 vaccine may be impaired in patients with rheumatic diseases, many areas remain to be clarified. There is an urgent need for real-world data on the immune response to the SARS-CoV-2 vaccine in patients with IMIDs treated with immunosuppressants and on the incidence and severity of SARS-CoV-2 infection in adequately vaccinated patients.Our main aim was to study antibody-mediated and cellular-mediated responses after SARS-CoV-2 vaccination in patients with IMIDs treated with different types of immunosuppressive drugs. Our secondary objectives were as follows: to describe the frequency and severity of SARS-CoV-2 infections after vaccination in patients with IMIDs, and to assess the B-cell compartment response after SARS-CoV-2 vaccination in patients treated with rituximab.We performed an ambispective observational study of patients with various IMIDs, namely, rheumatoid arthritis (RA), spondyloarthritis (SpA), psoriatic arthritis (PsA), and connective tissue disease (CTD). Patients were receiving b/tsDMARDs, as follows: TNF inhibitors (TNFi), rituximab, anti-interleukin 6 receptor (anti-IL6R) agents, and JAKi.Patients were recruited consecutively over 3 months, during which time they were invited to participate at their visits to the clinic (the month before and after the second and third doses of vaccine). The inclusion criteria were as follows: (i) IMIDs treated with b/tsDMARDs; and (ii) availability of laboratory tests 4–6 weeks after the second and/or third dose of the SARS-CoV-2 vaccine. The exclusion criteria were as follows: (i) IMIDs and not having received the second and/or third dose of the vaccine; and (ii) having received the vaccine more than 6 weeks previously.Demographic, clinical, and treatment data were collected from the electronic clinical records and the database used in the Complex Therapy Unit of the Rheumatology Department. The incidence and severity of SARS-CoV-2 infection after the 2nd and 3rd dose and data on disease flares were also recorded. These data on the incidence of infections and flares were obtained in the successive clinic visits until 6 months after the 3rd vaccine. In case of failing to attend it in person, they were contacted by telephone.Serum samples were obtained 4–6 weeks after the second or third dose to evaluate the humoral, cell-mediated, and functional response to the vaccine.The humoral response was assessed in all of the patients included: samples were available for 63 patients after the second and third doses, for 3 patients only after the second dose, and for 20 patients only after the third dose. In addition, 38 controls were selected to compare the humoral response with that of the patients after the second dose.The humoral response was assessed using fluoroenzyme immunoassay  with serum samples. The lower limit of detection was 0.7 IU/ml, and the positive cut-off was set at 10 IU/ml. The upper limit of detection was 204 IU/ml. Data from IMIDs patients were compared with those from the 38 healthy controls analyzed 4 weeks after the second vaccine dose. In this cohort, samples above 204 IU/ml were subsequently diluted to obtain a final antibody concentration.The cellular response was evaluated only in patients with a poor humoral response, which was defined as IgG antibody levels against SARS-Cov2 < 100 IU/ml, using a QuantiFERON SARS-CoV-2 (QTF-SARS-COV-2) Starter Pack (Qiagen). A cut-off value of 0.15 IU/ml was applied to differentiate between positive and negative cell-mediated immune responses.+ B cells in peripheral blood. The B-cell phenotype in RA patients was compared with our 62 previously characterized HC cohort .Some RA patients had normal absolute total CD19ols Fig. A. In add++ CD21het CD24++ immature/transitional B lymphocytes RA patients  and in the other 2 more differentiated immature/transitional B-cell subsets: CD5+ CD38+/++ CD21het CD24++ (14.59 vs 5.60 cells/µl in HD) and CD5+ CD38het CD21+ CD24+ (30.14 vs 16.90 cells/µl in HD)  . The frequency of SARS-COV-2 infection was similar in each treatment group: 58% 19/45) for TNFi, 55% (14/31) for rituximab, 60% (2/5) for anti-IL6R, and 60% (2/5) for JAKi . Disease activity was controlled by adjusting treatment with corticosteroids, although 1 patient required a change in biological treatment.This study provides insight into the clinical relevance of immune response to SARS-COV-2 vaccination in patients with IMIDs treated with immunosupressants. In our cohort, most of patients (86%) with IMIDs treated with b/tsDMARDs had detectable humoral responses. However, anti-SARS-COV-2 Ab levels were lower in patients than in controls, with rituximab being associated with a higher risk of an absent humoral response (OR: 18.5), as expected. On the other hand, we found that up to 37% of patients with a poor humoral response may have a cellular response to the vaccine, being more frequently detected in patients treated with rituximab (91%). Another important issue is that the presence of CD19+ B cells in peripheral blood is associated with the time since the last rituximab infusion. The study of the pre-GC B cell compartment showed that the B cell regeneration pattern in patients is mainly at the expense of immature cells.There is substantial variation in the immune response to vaccination between individuals. Factors that influence humoral and cellular vaccine responses include intrinsic host factors such as age, sex, genetics, and comorbidities, perinatal factors such as gestational age, birth weight, feeding method, and maternal factors, as well as extrinsic factors like preexisting immunity, microbiota, infections, and antibiotics . AdditioMoreover, factors related with vaccine  and administration  are also important , 27. An The seroconversion rate in our cohort of patients with IMIDs was very high 86%). However, anti-SARS-CoV-2 Ab levels were lower in patients with IMIDs treated with b/tsDMARDs than in controls. These data are consistent with previous publications 6%. Howev. These fAnother interesting point to describe in detail is the cellular response that patients with IMIDs can generate against the SARS-COV-2 vaccine. Lledó et al. found that cellular immune response against SARS-CoV-2 was similar between patients with IMIDs (53) and controls (61) . They al− B-cells have been described in the naïve B-cell compartment and in the memory B cell (MBC) compartment [− CD24− mature naïve B cells. Although we have not evaluated the B-cell phenotype prior to the rituximab treatment, it could be presumed that the regeneration of B-cell compartment, with accumulation of CD21− B-cells, is similar to the preexisting one which is the result of the chronic stimulation occurred in the pathogenesis of RA.Delving into the mechanism of rituximab, CD20 is expressed on circulating peripheral blood B cells, but not on bone marrow stem cells or plasma cells . Therefopartment . Here, t+ MBC and switched IgMD+ MBC, as also previously reported in systemic lupus erythematous (SLE) patients treated with rituximab [In concordance with delayed MBC compartment reconstitution, seven patients with RA presented severely reduced unswitched IgMDituximab . ConsideA very reassuring fact is that in our cohort there is no difference in the frequency of infections between patients and controls after vaccination and they were mostly mild cases. The infections requiring hospitalization after vaccination were scarce and they only occurred in patients who were treated with rituximab. These results support the importance of promoting an adequate vaccination strategy as stated in the EULAR recommendations for vaccination in patients with IMIDs . HoweverFollowing the recommendations from the Ministry of Health, all patients received additional COVID-19 vaccine boosters . It is evident that vaccination against SARS-COV-2 has decreased the frequency and severity of infections. In fact, our data show that the majority of patients presented mild infections after vaccination like those observed in the general population. However, an interesting point observed in this work is that the antibody titers were significantly lower after the 3rd dose than after the initial regimen that included the 1st and 2nd doses. This fact reflects how the antibody response diminishes over time and may be related to the dose used and/or the timing of the booster. Therefore, the frequency and number of boosters needed for these patients are unclear. In this sense, efforts should be made to move towards more personalized strategies, establishing profiles of patients at higher risk based on scientific evidence.” In this sense, efforts should be made to move towards more personalized strategies, establishing profiles of patients at higher risk based on scientific evidence.Some limitations were observed in the present study. First, the limited number of patients in the groups of patients treated with anti-IL6R and JAKi. Second, some differences in age and BMI between controls and patients. However, these age differences were only observed in patients with RA. Regarding BMI, recent data do not seem to find a relationship between BMI and response to the SARS-COV-2 vaccine. Another aspect to consider is that the cellular response was only studied in patients with a poor humoral response, so we cannot compare the cellular response between patients with a good humoral response or not.On brief, the humoral immune response to the SARS-COV-2 vaccine is poorer in patients than in healthy controls, especially in those treated with rituximab. However, in patients with rituximab, a cellular-mediated immune response can be detected despite having a poor humoral response. Finally, severe infections in vaccinated patients with IMIDs are rare, mainly observed in patients treated with rituximab.In conclusion, a scheduled delay of the rituximab infusion to ensure a better humoral response to the vaccine would be recommended in patients with IMID.Additional file 1: Table S1. Comparison of characteristics between controls and patients classified by diagnosis. Table S2. SARS-CoV-2 vaccination patterns in patients and controls. Table S3. Comparison of patients with low anti-SARS-Cov-2 antibody levels (< 100 IU/ml) in each therapy group. Figure S1. Distribution of B-cell subsets by degree of maturation, including immature cells , naïve cells , and memory B cells (MBC) in peripheral blood. Figure S2. Humoral immune response after the second vaccine dose in patients with IMIDs. Comparison of the second vaccine dose between controls and patients. Figure S3. Proportion of patients with SARS-CoV-2 infections in each therapy group. This graph shows the proportion of patients who developed SARS-CoV-2 infection and its severity after the second dose of vaccine according with each treatment.